荧光光谱法研究利血平的氧化作用及其分析应用

利血平自身有弱荧光,氧化后可形成较强荧光物质。本文系统地研究了稀ＨＮＯ３对利血平的氧化作用,提出了高灵敏度测定利血平含量的荧光光度分析新方法。结果表明,荧光强度和利血平的浓度在０．０２－０．４μｇ／ｍＬ范围内呈良好的线性关系,方法的检出限为０．００２μｇ／ｍＬ（６．７３＊１０＾－９ｍｏｌ／Ｌ）。该法简便,快速、灵敏度高,用于注射液中利血平含量的测定,结果令人满意。     

利血平与玫瑰精B的高灵敏显色反应及分析应用

研究了利血平与玫瑰精B的显色反应,建立了测定利血平的高灵敏分光光度法。在酸性条件下,利血平的水解产物和玫瑰精B形成具有正、负吸收峰的红色离子缔合物,最大正吸收波长位于490 nm,最大负吸收波长位于520 nm,表观摩尔吸光系数(ε)分别为1.20×105L.mol-1.cm-1(正吸收)和1.83×105L.mol-1.cm-1(负吸收),利血平在0～5.0μg/mL范围内遵从比尔定律。若采用正、负峰叠加测定,灵敏度可达3.00×105L.mol-1.cm-1。探讨了适宜的反应条件、主要分析特性及方法的精密度和可靠性。该法可用于市售利血平注射液中利血平含量的测定。     

Flow—injection Chemiluminescence Determination of Reserpine in Medicine and Biological Fluids with Controlled—Reagent—Release Technology

A sensitive and rapid chemiluminescence(CL) flow injection with controlled-reagent-release technology for the determination of reserpine was proposed.The Cl reagents,luminol and dichromate,uses in this sensor,were all immobilized on anion-exchange resin.Through injection of 100μl of water,the reagents on the anion-ex-change resin column were eluted and in the presence of reserpine ,the CL intensity was decreased,by which reserpine could be sensed.Reserpine was quantified by measuring the decrement of CL intensity,which was observed linear with the logrithm of reserpine concentration in the rage of 1.0-500.0ng/mL,and the limit of detection was 0.4ng/mL(3σ)with a relative standard deviation of less than 3.0?The proposed procedure was applied in the assay of reserpine in pharmaceutical preparation and biological fhuids without any pre-treatment process and with sampling frequencies of 72 times per hour.     

High Performance Liquid Chromatographic Determination of Diuretic-Antihypertensive Combination Products. II. Polythiazide and Reserpine

Abstract

A high performance liquid chromatographic method for the simultaneous determination of polythiazide and reserpine in tablets is described. Polythiazide, reserpine, and vanillin (an excipient) are separated isocratically on an octadecylsilane column using a methanol-water-acetic acid (55:44:1) mobile phase. The column effluent was monitored by ultraviolet ab-sorbance at 254 nm. Recoveries from synthetic formulations were 100.2 ± 1.7% for polythiazide and 98.7 ± 1.1% for reserpine.     

Chemiluminometric determination of reserpine and related alkaloids

The determination of the alkaloids reserpine, rescinnamine and yohimbine based on a chemiluminogenic reaction with potassium permanganate in the presence of polyphosphoric acid is described. The investigation was carried out using a batch and a flow injection chemiluminometer. Both approaches were accurate and precise, allowing the measurement of reserpine within the ranges 0.100-3.00 and 0.050-3.00 micrograms ml-1 with RSD values for 1.00 microgram ml-1 of 1.91 and 0.33% (n = 8) with the batch and the flow injection manifold, respectively. The procedure was successfully applied to formulations after extraction of reserpine with chloroform, with recoveries from commercial formulations within the range 95.2-99.0%.     

Fluorometric determination of reserpine in dosage forms

A rapid and highly sensitive fluorometric procedure has been developed for the routine determination of reserpine, in bulk and in dosage forms. The method is based on the fluorescence induced by oxidation of reserpine with hexa-amminecobalt(III) tricarbonatocobaltate in aqueous acetic acid. The oxidation product exhibits a greenish yellow fluorescence with its emission maximum at around 420 nm. The fluorescence intensity is a linear function of reserpine concentration over the range 0.01-0.24 mug/ml in the solution finally measured. The advantages and disadvantages of the proposed method are discussed, and its applicability to different formulations is demonstrated.     

Spectrophotometric determination of Rauwolfia alkaloids: estimation of reserpine in pharmaceuticals.

A simple, sensitive and economically viable spectrophotometric method for the determination of some Rauwolfia alkaloids (ajmaline, ajmalicine, reserpine and yohimbine-HCl) has been developed. The method involves the oxidation of Rauwolfia alkaloids by iron(III) and subsequent complexation of iron(II) with 1,10-phenanthroline, forming a red-colored complex having the maximum absorbance at 510 nm. The method is applied to the determination of reserpine in tablets of pharmaceutical formulations. The common excipients do not interfere with the proposed method. A statistical comparison of these results with those of a reported method shows good agreement and indicates no significant difference in the precision.     

毛细管电泳法测定非水溶性药物——复方降压片

在区带电泳中,应用在背景电解质中加入乙腈的方法对复方降压片的两种主要成分利血平、双氢氯噻嗪进行了分离和定量测定。５次重复测定的相对标准偏差小于３％,检出限分别为０．２６９ｍｇ／Ｌ和１．８４ｍｇ／Ｌ,并对ｐＨ值、磷酸盐浓度及乙腈用量进行了考察。     

Electrospray-ionization driven by dielectric polarization

A non-conductive piezo ceramic plate has been used to induce an electric field to generate an electrospray as ionization method for mass spectrometric determination. This technique decreases the risk of undesired discharges, induced by high electric currents. The applicability of the technique is demonstrated and compared with a commercial electrospray for mass spectrometric determination of reserpine and myoglobin.     

A new gas chromatographic method for the estimation of reserpine and rescinnamine.

Under the conditions described for alkaline hydrolysis of reserpine and rescinnamine in absolute and aqueous methanol, and after esterification (with diazomethane) of the resulting acid fraction, methyl 3,4,5-trimethoxybenzoate was quantitatively recovered, whereas methyl trans-3,4,5-trimethoxycinnamate, in normal lighting conditions, was either partly isomerized to methyl cis-trimethoxycinnamate or formed an adduct with a molecule of methanol, yielding methyl 3-methoxy-3-(3,4,5-trimethoxyphenyl)propionate. The structures of the products were established by synthesis, nuclear magnetic resonance studies and mass spectrometry. This investigation of the hydrolytic conditions allowed a reliable and rapid gas chromatographic determination of reserpine and/or rescinnamine in amounts down to 500 and 2000 mug, respectively, to be devised.     

Sensitive Adsorption Stripping Voltammetric Determination of Reserpine by a Glassy Carbon Electrode Modified with Multi-Wall Carbon Nanotubes

Adsorption stripping voltammetry, a very sensitive electroanalytical method, was employed to determine reserpine, a kind of anti-hypertensive drug. In 0.1M phosphate buffer with a pH of 6.0, reserpine was accumulated at a multi-wall carbon nanotubes (MWNT)-modified glassy carbon electrode (GCE) surface under the condition of open-circuit. In the following anodic sweep from 0.20 to 1.00V, reserpine, adsorbed at the MWNT-modified GCE surface, was oxidized and yielded a sensitive oxidation peak at 0.64V. Due to its unique structure and extraordinary properties, MWNT shows a ten times higher accumulation efficiency toward reserpine, compared with a bare GCE. Hence, the amount of reserpine at the MWNT-modified GCE surface increases significantly, and finally the oxidation peak current improves greatly. The experimental conditions, such as supporting electrolyte, pH value, the amount of MWNT-DHP suspension, accumulation time and scan rate, were optimized for the measurement of reserpine, and a sensitive electroanalytical method was proposed for reserpine determination. The oxidation peak current varies linearly with the concentration of reserpine over the range of 2×10^–8 to 1×10^–5M, and the detection limit is 7.5×10^–9M after 4min open-circuit accumulation. The relative standard deviation at 1×10^–6M reserpine was about 4.7% (n=7), indicating excellent reproducibility. This new method was successfully demonstrated with reserpine injections and tablets.     

Simultaneous Determination of Hydrochlorothiazide and Reserpine in Human Urine by LC with a Simple Pre-Treatment

A simple, selective and sensitive reversed-phase high performance liquid chromatography method for simultaneous analysis of hydrochlorothiazide and reserpine in human urine was developed and subjected to primary pharmacokinetic study. After a simple protein precipitation using methanol and extraction with ethyl acetate, the analytes were separated on an Elite C(18) column at a flow rate of 0.8 mL min(-1). The mobile phase was composed of acetonitrile (A) and 0.2% ammonium chloride solution (B) for a gradient elution starting at A:B at 30:70, v/v for 0~6 min, linearly raising the percent of A from 30% to 50% (6~9 min) and ending at 50:50, v/v (9~25 min). The standard curves were linear over the range of 0.05-20 μg mL(-1) for hydrochlorothiazide and 0.02-5.0 μg mL(-1) for reserpine, respectively (r > 0.999). The limit of detection (LOD) and the limit of quantification (LOQ) were 5.5 ng mL(-1) and 18.2 ng mL(-1) for hydrochlorothiazide, and 7.1 ng mL(-1) and 23.6 ng mL(-1) for reserpine, respectively. The recoveries for both analytes were above 89.0±1.35%. The intra-day and inter-day precision for hydrochlorothiazide were less than 1.91% and 1.38%, and those for reserpine were below 1.61% and 2.64%, respectively. The method indicated good performance in terms of specificity, linearity, detection and quantification limits, precision and accuracy, and it was employed successfully for the simultaneous determination of hydrochlorothiazide and reserpine in human urine samples.     

Determination of individual proton affinities of reserpine from its UV-vis and charge-transfer spectra

Proton affinities of the two N atoms of reserpine (methyl-11,17alpha-dimethoxy-18beta-[(3,4,5-trimethoxybenzoyl)oxy]-3beta,20alpha-yohimban-16beta-carboxylate) have been determined in two ways from the pH-dependent variation of the UV-vis absorption spectra (i) of reserpine itself and (ii) of the charge-transfer (CT) spectra of its complexes with o-chloranil, p-chloranil, and DDQ in aqueous medium (containing 0.1% ethanol v/v). For the second method, the CT absorption bands of the complexes were determined, their formation constants were estimated by a modified Benesi-Hildebrand equation, and variation of CT absorption spectra with a change in pH was noted. A necessary working formula for the second method was derived and utilized with the experimental data. The pKa values obtained by the two methods are well in agreement with each other within the limits of experimental error. To our knowledge, so far, this is the first report on determination of pKa from charge-transfer complex formation in aqueous solution using simple absorption spectroscopy in the UV-vis region. The results obtained were further checked by noting the variation of fluorescence intensity of reserpine upon addition of o-chloranil, acid, and base, and almost complete agreement with the absorption spectrometric result was observed.     

Determination of Reserpine in Plasma Using High-Performance Liquid Chromatography with Fluorescence Detection

Abstract

A high-performance liquid chromatographic method for determining reserpine in plasma has been developed. The procedure involves extraction of reserpine from buffered plasma into benzene, oxidation of reserpine to a fluorophor by treatment with vanadium pentoxide in phosphoric acid, and chromatographic separation of the reserpine fluorophor on an octadecylsilane column by ion-pairing with heptanesulfonate ions. Fluorescence monitoring of the column effluent provides high sensitivity of detection and increases the specificity of the procedure. A detection limit of approximately 100 pg of reserpine per ml of plasma was obtained following analysis of 2 ml samples. Analysis of a number of samples demonstrated the applicability of this method in confirming the presence of reserpine in equine plasma specimens collected at various horse shows and in evaluating the pharmacokinetic behavior of reserpine following intramuscular administration to horses.     

Flow injection spectrofluorimetric determination of reserpine in tablets by on-line acetone sensitized photochemical reaction

On-line photochemical reaction of reserpine in the presence of acetone was investigated. Acetone was found to speed up the on-line photochemical conversion of reserpine into an intensively fluorescent compound. Not only reaction acidity but also the acetate buffer concentration affected the on-line photochemical induced fluorescence signal. Based on the observation an automated flow injection photochemical fluorimetric approach was developed. An injected sample zone was carried by a water stream to be merged with a acetate buffer (pH 3.4) solution containing 0.02% acetone in a knotted PTFE reactor (KR), which was freely coiled around a 6-W low pressure mercury lamp. While passing the KR, reserpine was transformed into an intensively fluorescent compound. It was on-line detected in a flow-through cell at the emission wavelength of 490 nm and excitation wavelength of 386 nm. At optimized conditions, a detection limit 0.45 mug l(-1) was achieved at a sampling rate of 90 h(-1). Eleven determinations of a 0.5 mg l(-1) reserpine standard solution gave a R.S.D. of 0.3%. The linear dynamic range of reserpine calibration curve was 0.01-0.75 mg l(-1). The proposed method was applied to assay the reserpine content in tablets and to monitor the dissolution profile of reserpine tablets. Satisfactory results were obtained for both the assays and dissolution studies.     

Determination of Indole Alkaloids by High-Performance Liquid Chromatography with Resonance Rayleigh Scattering Detection

A novel resonance Rayleigh scattering (RRS) detection approach combined with high–performance liquid chromatography (HPLC) was developed for the determination of reserpine, deserpidine, and ajmalicine. The resonance Rayleigh scattering signal increased when the analytes were bound to diiodofluorescein in Britton–Robinson buffer (pH 4.2). Separation was performed using a C18 column with a mobile phase consisting of acetonitrile–5 millimolar ammonium acetate buffer at pH 4.2 (44:56, volume by volume). Resonance Rayleigh scattering was measured at excitation and emission wavelengths of 320 nanometers. The experimental parameters affecting the separation and the scattering intensity were carefully optimized. Possible mechanisms for the resonance Rayleigh scattering enhancement of the indole alkaloid–diiodofluorescein system were explored by scanning electron microscopy, nuclear magnetic resonance, and ultraviolet-visible absorption spectroscopy. The method was employed for the determination of the analytes in human urine and pharmaceutical tablets.     

Determination of sub-ppb reserpine by an optosensing device based on photochemically induced fluorescence

A flow injection–solid-phase spectroscopy (FI-SPS) system implemented with photochemically induced fluorescence (PIF) is described for the rapid and very sensitive determination of reserpine in biological fluids and pharmaceutical formulations. An intensively fluorescent photoproduct is in-line generated, retained on C₁₈ silica gel in the detection area and monitored at 394/489 nm (λ _ex/λ _em). After the establishment of the appropriate working variables, the system is calibrated at two different injection volumes, 100 and 800 μL, achieving detection limits of 0.33 and 0.05 ng mL⁻¹, respectively. The RSD for reserpine at 2 ng mL⁻¹ (800 μL) was 1.5% (n = 10). The sampling rates were 46 and 43 h⁻¹ for each injection volume, respectively. The potential interference of some common species coexisting with reserpine in the analysed samples was also studied. The procedure was successfully applied to commercial formulations, urine and serum without any previous treatment of samples. Recoveries ranged from 94.9 to 100.2%.     

用大孔吸附树脂分离利血平

以利血平的吸附量和解吸率为指标,筛选大孔吸附树脂.研究吸附和解吸的优化条件,并考察选定树脂的吸附等温线、吸附动力学、吸附和解吸性能.结果表明,将催吐萝芙木根粉浸提液蒸去乙醇且不调pH(pH 1)进行吸附,HZ-818型大孔吸附树脂对利血平的吸附量可达到9.34mg/mL.使用工业乙醇-水(80:20,pH 1.0)为解吸剂,解吸率可达99.3%.该树脂的吸附符合Langmuir吸附等温方程.吸附前期,吸附速度较快,以后速度减慢.HZ-818型树脂对利血平的吸附量大,解吸率高,通过大孔树脂吸附和解吸,利血平浓度提高50倍以上,适宜于工业化生产.     

Characterization of dielectric barrier electrospray ionization for mass spectrometric detection

A novel electrospray interface is presented which induces an electric field by dielectric polarization through a non-conductive barrier. Therefore, a square-wave high-voltage signal is applied. This technique allows mass spectrometric measurements in the positive as well as in the negative mass spectrometry mode without changing the polarity of the potential applied, and it decreases the risk of undesired discharges, induced by high electric currents. The applicability of this technique is demonstrated by mass spectrometric determination of reserpine.     

Practical Microbore Column HPLC: System Development and Drug Applications

Abstract

Commercially available packed microbore columns have been used to demonstrate the potential of microbore high performance liquid chromatography (micro-LC) systems. The necessity of optimizing the chromatographic system for micro-LC is demonstrated using sulfa drugs and antibiotic standards, and some of the advantages of micro-LC such as high mass sensitivity are shown. Finally micro-LC has been used for the determination of two drugs, reserpine and trichlormethiazide in biological fluids.